Real-Time Cell Cycle Imaging in a 3D Cell Culture Model of Melanoma

Spoerri, Loredana; Beaumont, Kimberley A.; Anfosso, A.; Haass, Nikolas K.

doi:10.1007/978-1-4939-7021-6_29

Cited by 50 publications

(98 citation statements)

References 21 publications

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“…a , Supporting Information Figures S2B‐C). To study invasion, we utilized a 3D melanoma spheroid model that mimics in vivo tumor architecture and microenvironment . RAB27A ‐knockdown (KD) in these RAB27A‐high cell lines resulted in strongly reduced invasion into the collagen matrix using three effective RAB27A shRNAs (sh#1, sh#2, sh#4; Figs.…”

Section: Resultsmentioning

confidence: 99%

“…The human melanoma cell lines C8161, 1205Lu, WM793, WM983B, WM983C, WM164 and 451Lu were genotypically characterized and grown as described . All human melanoma cell lines and HEK293T were authenticated by STR fingerprinting (Molecular Genetics facility, Garvan Institute of Medical Research, Darlinghurst, NSW, Australia).…”

Section: Methodsmentioning

confidence: 99%

“…B16‐F10 melanoma cells were obtained from ATCC. WM164 cells stably expressing Fluorescence Ubiquitination Cell Cycle Indicator (FUCCI) were created as previously described . Cells were tested for mycoplasma using DAPI staining and high‐magnification microscopy, as well as PCR at Cell Bank Australia.…”

Section: Methodsmentioning

confidence: 99%

“…Melanoma spheroids were prepared as described previously . At least three spheroids were implanted into collagen per condition, experiments were repeated at least three times.…”

Section: Methodsmentioning

confidence: 99%

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RAB27A promotes melanoma cell invasion and metastasis via regulation of pro‐invasive exosomes

Guo

Lui

Lai

et al. 2019

Intl Journal of Cancer

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Despite recent advances in targeted and immune‐based therapies, advanced stage melanoma remains a clinical challenge with a poor prognosis. Understanding the genes and cellular processes that drive progression and metastasis is critical for identifying new therapeutic strategies. Here, we found that the GTPase RAB27A was overexpressed in a subset of melanomas, which correlated with poor patient survival. Loss of RAB27A expression in melanoma cell lines inhibited 3D spheroid invasion and cell motility in vitro, and spontaneous metastasis in vivo. The reduced invasion phenotype was rescued by RAB27A‐replete exosomes, but not RAB27A‐knockdown exosomes, indicating that RAB27A is responsible for the generation of pro‐invasive exosomes. Furthermore, while RAB27A loss did not alter the number of exosomes secreted, it did change exosome size and altered the composition and abundance of exosomal proteins, some of which are known to regulate cancer cell movement. Our data suggest that RAB27A promotes the biogenesis of a distinct pro‐invasive exosome population. These findings support RAB27A as a key cancer regulator, as well as a potential prognostic marker and therapeutic target in melanoma.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…Melanoma spheroids were prepared as described previously . At least three spheroids were implanted into collagen per condition, experiments were repeated at least three times.…”

Section: Methodsmentioning

confidence: 99%

See 2 more Smart Citations

RAB27A promotes melanoma cell invasion and metastasis via regulation of pro‐invasive exosomes

Guo

Lui

Lai

et al. 2019

Intl Journal of Cancer

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“…The acidic pH of collagen was neutralized by including 7.5% (w/v) sodium bicarbonate in the gel mixture . After solidification of the mixture of collagen gel and the spheroids, a culture medium consisting of DMEM+10% FBS, 2 mM l ‐glutamine, and 1% penicillin–streptomycin (all from Gibco) was added to cover the gel mixture.…”

Section: Methodsmentioning

confidence: 99%

miR‐16 enhances miR‐302/367‐induced reprogramming and tumor suppression in breast cancer cells

2020

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Overexpression of either miR‐302 or miR‐302/367 cluster induces reprogramming of cancer cells and exerts tumor‐suppressive effects by induction of mesenchymal‐to‐epithelial transition, apoptosis and a less proliferative capacity. Several reports have described miR‐16 as a tumor suppressor microRNA (miRNA). Here, we studied the impact of exogenous induction of miR‐16 in MDA‐MB‐231 and SK‐BR‐3 breast cancer cells following overexpression of miR‐302/367 cluster and investigated whether transfection of these cells by a mature miR‐16 mimic could affect the reprogramming state of the cells and their tumorigenicity. miR‐16 enhanced the expression levels of OCT4A, SOX2, and NANOG, generally known as transcription or pluripotency factors, and suppressed proliferation and invasiveness of these cells. Meanwhile, inhibition of miR‐16 counteracted both the reprogramming effect and the antitumor function of miR‐302/367 in the breast cancer cells. Current results indicate that miR‐16 can work as an adjuvant to improve both cancer cell reprogramming and tumor‐suppressive function of miR‐302/367 cluster in MDA‐MB‐231 and SK‐BR‐3 cells, while its inhibition counteracts all of these effects. Combined application of miRNAs that share some common targets in cancer cell signaling pathways may provide new approaches for repression of multiple hallmarks of cancer.

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Magnolol induces cell death through PI3K/Akt‐mediated epigenetic modifications boosting treatment of BRAF‐ and NRAS‐mutant melanoma

et al. 2019

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Most BRAF‐mutant melanoma patients experience a fulminate relapse after several months of treatment with BRAF/MEK inhibitors. To improve therapeutic efficacy, natural plant‐derived compounds might be considered as potent additives. Here, we show that magnolol, a constituent of Magnolia officinalis, induced G1 arrest, apoptosis and cell death in BRAF‐ and NRAS‐mutant melanoma cells at low concentration, with no effect in BRAF‐ and NRAS wild‐type melanoma cells and human keratinocytes. This was confirmed in a 3D spheroid model. The apoptosis‐inducing effect of magnolol was completely rescued by activating Akt suggesting a mechanism relying primarily on Akt signaling. Magnolol significantly downregulated the PI3K/Akt pathway which led to a global decrease of the active histone mark H3K4me3. Alongside, the repressive histone mark H3K9me3 was increased as a response to DNA damage. Magnolol‐induced alterations of histone modifications are reversible upon activation of the Akt pathway. Magnolol‐induced a synergistic effect in combination with either BRAF/MEK inhibitors dabrafenib/trametinib or docetaxel at a lower concentration than usually applied in melanoma patients. Combination of magnolol with targeted therapy or chemotherapy also led to analogous effects on histone marks, which was rescued by Akt pathway activation. Our study revealed a novel epigenetic mechanism of magnolol‐induced cell death in melanoma. Magnolol might therefore be a clinically useful addition to BRAF/MEK inhibitors with enhanced efficacy delaying or preventing disease recurrence.

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Real-Time Cell Cycle Imaging in a 3D Cell Culture Model of Melanoma

Cited by 50 publications

References 21 publications

RAB27A promotes melanoma cell invasion and metastasis via regulation of pro‐invasive exosomes

RAB27A promotes melanoma cell invasion and metastasis via regulation of pro‐invasive exosomes

miR‐16 enhances miR‐302/367‐induced reprogramming and tumor suppression in breast cancer cells

Magnolol induces cell death through PI3K/Akt‐mediated epigenetic modifications boosting treatment of BRAF‐ and NRAS‐mutant melanoma

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